This invention relates to the field of separation science and analytical biochemistry using gas phase ion spectrometry, in particular mass spectrometry. Typically, analysis of biological samples by mass spectrometry involves the desorption and ionization of a small sample of material using an ionization source, such as a laser. The material is desorbed into a gas or vapor phase by the ionization source, and in the process, some of the individual molecules are ionized. Then the ionized molecules can be dispersed by a mass analyzer and detected by a detector. For example, in a time-of-flight mass analyzer, the positively charged ionized molecules are accelerated through a short high voltage field and let fly (drift) into a high vacuum chamber, at the far end of which they strike a sensitive detector surface. Since the time-of-flight is a function of the mass of the ionized molecule, the elapsed time between ionization and impact can be used to identify the presence or absence of molecules of specific mass.
Desorption mass spectrometry had been around for some time. However, it was difficult to determine molecular weights of large intact biopolymers, such as proteins and nucleic acids, because they were fragmented (destroyed) upon desorption. This problem was overcome by using a chemical matrix. In matrix-assisted laser desorption/ionization (MALDI), the analyte solution is mixed with a matrix solution (e.g., a very large molar excess of an acidic, UV absorbing matrix solution). The mixture is allowed to crystallize after being deposited on an inert probe surface, trapping the analyte within the crystals. The matrix is selected to absorb the laser energy and apparently impart it to the analyte, resulting in desorption and ionization. See, U.S. Pat. No. 5,118,937 (Hillenkamp et al.), and U.S. Pat. No. 5,045,694 (Beavis & Chait).
Recently, surface-enhanced laser desorption/ionization (SELDI) was developed which is a significant advance over MALDI. In SELDI, the probe surface is an active participant in the desorption process. One version of SELDI uses a probe with a surface chemistry that selectively captures analytes of interest. For example, the probe surface chemistry can comprise binding functionalities based on oxygen-dependent, carbon-dependent, sulfur-dependent, and/or nitrogen-dependent means of covalent or noncovalent immobilization of analytes. The surface chemistry of a probe allows the bound analytes to be retained and unbound materials to be washed away. Subsequently, analytes bound to the probe surface can be desorbed and analyzed using mass spectrometry. This method allows samples to be desorbed and analyzed directly without any intermediate steps of sample preparation, such as sample labeling or purification. Therefore, SELDI provides a single, integrated operating system for the direct detection of analytes. SELDI and its modified versions are described in U.S. Pat. No. 5,719,060 (Hutchens & Yip) and WO98/59361 (Hutchens & Yip).
The desorption methods described above have unlimited applications in the field of separation science and analytical biochemistry. For example, cell surface or soluble receptors can be attached to the probe surface to screen for ligands. Bound ligands can then be analyzed by desorption and ionization. Nucleic acid molecules can also be attached to the probe surface to capture biomolecules from complex solutions. Biomolecules, which are bound to the nucleic acid, can then be isolated and analyzed by desorption and ionization. Furthermore, antibodies attached to the probe surface can be used to capture and identify specific antigens. The antigens which are specifically bound to the antibody can then be isolated and analyzed by desorption and ionization.
While the probes described above provide a great tool in the field of separation science and analytical biochemistry, it would be desirable to develop a probe having a surface chemistry that provides an increased capacity and sensitivity. When the amount of sample available for analysis is very small and limited, it would be desirable to have a desorption system having an increased sensitivity of detection. Furthermore, it would be also desirable to develop a probe capable of providing consistent mass resolution and intensities of bound analytes on the probe.